Activation of Nonspecific Cytotoxic Cells with a Multiple Antigenic Peptide: Specificity and Requirements for Receptor Crosslinkage

Nonspecific cytotoxic cells (NCC) of teleost fish recognize a conserved antigenic determinant found on the protozoanTetrahymena pyriformisand on many different tumor target cells. This determinant is located on a 46-kDaTetrahymenaprotein referred to as natural killer target antigen (NKTag). The NKTag cognate sequence recognized by NCC is composed of seven to nine amino acids. In the present study, synthetic peptides of the cognate NKTag determinant were prepared as multiple antigenic peptides (MAP). Immobilized MAP activated NCC lysis of IM-9 target cells in the absence of antigen presenting cells or exogenous added cytokines. NCC binding to immobilized MAP produced two- to fivefold increased lysis of IM-9, U937, and HL-60 target cells compared to scrambled control MAP (composed of the same amino acids only in random sequence). NCC receptor crosslinkage was required for activation. Immobilized monomeric homologous cognate peptide did not activate increased NCC lysis of IM-9 target cells; however, NCC preincubated with soluble homologous monomer inhibited MAP activation of lysis. Ligand activation of NCC was antigen specific. Binding of the immobilized ligand with anti-MAP mab 22A12 prior to addition of NCC blocked activation. Mab 22A12 also inhibited NCC lysis of IM-9 target cells. A possible mechanism of NCC activation was determined. Binding of NCC to immobilized MAP produced significantly increased membrane expression of a putative receptor as defined by mab 5C6 binding. These studies demonstrate that activation of NCC by a target cell antigenic determinant depends on crosslinkage of an NCC “receptor” by polymeric repetitive sequences, a soluble or fixed monomer cannot activate but can inhibit activation, and MAP binding initiates increased expression of a putative NCC receptor protein.